Explanation Diabetes and obesity are emerging worldwide health problems. New prevention and treatment options for both conditions could be based on strategies to dampen or inhibit nutrient absorption. Similar strategies are the basis of agents currently used clinically to inhibit fat absorption, cholesterol absorption, and intestinal catabolism of complex carbohydrates. A new class of agents that delayed or inhibited glucose absorption could have substantial impact in managing diabetes and obesity. Emerging evidence indicates that apical, or luminal facing, GLUT2 is a major pathway of sugar absorption, and is therefore an attractive target of such potential agents. Flavonoids are polyphenols that are widely distributed in foods, especially fruits and vegetables. Nutritive functions of flavonoids are unknown. Quercetin is a commonly ingested flavonoid, and 20-100 mg daily is ingested by dietary intake. Peak plasma concentrations of flavonoids such as quercetin do not exceed 1-2 umoles per liter after ingestion, but intestinal luminal concentrations are believed to be approximately 50 fold higher. Based on these high intraluminal concentrations, we proposed that a novel action of intraluminal flavonoids may be to dampen, re-distribute, or frankly inhibit intestinal absorption of candidate nutrients. Flavonoids either as food components or co-administered with foods could potentially have these actions, and such actions would not require flavonoids themselves to be absorbed. Partial support for this proposal was provided by data showing that some flavonoids found in foods inhibited vitamin C and glucose intestinal transport and absorption. The findings suggested that quercetin, the dominant flavonoid ingested by humans, may modulate glucose intestinal absorption by the sodium-independent facilitative glucose transporter GLUT2. Although these findings are promising, a number of uncertainties remain. Some investigators suggested that flavonoids decreased glucose uptake by a sodium-dependent pathway, via the sodium-dependent glucose transporter SGLT1. This conclusion was based on experiments using intestinal cells, brush border membrane vesicles, or Xenopus laevis oocytes expressing SGLT1. In cell and vesicle preparations it is difficult to distinguish which transporter(s) are inhibited, and the concentrations of substrates and flavonoids used in all of these experiments were not relevant to in vivo conditions. In systems where SGLT1 was either overexpressed in cells or in Xenopus oocytes, flavonoid effects on glucose transport were either modest or not directly tested. Other investigators suggested that flavonoids could be non-specific inhibitors, based on their behavior in isolated enzyme systems. It is uncertain which intestinal glucose transporters are inhibited by different flavonoids, which flavonoids are the most potent inhibitors of glucose transport, whether there is selectivity for inhibition of transport of different substrates, whether flavonoids must first be deglycosylated or transported for inhibition to occur, and whether the appropriate transporters are in the same location as the inhibitory concentrations of flavonoids. Addressing these issues would provide a clear data base of flavonoid action that could serve as the foundation of a pilot clinical study of flavonoid effects on sugar absorption. To test transporter specificity, we studied oocytes that were injected with cRNAs to express specific intestinal sugar transporters, and incubated these oocytes with a variety of flavonoids to determine potency. To learn whether flavonoid inhibition of sugar transport was specific for glucose transporters expressed only in oocytes, cell systems were also studied. Flavonoids, flavonoid concentrations, and sugar substrate concentrations were all selected to have in vivo relevance. We showed that several flavonoids were potent inhibitors of GLUT2-mediated glucose and fructose transport but had no effect on other major intestinal sugar transporters; that flavonoid structure and glycosylation affected inhibition; that GLUT2 overexpressed in cells and present in an intestinal cell model was inhibited by the relevant flavonoids; that GLUT2 was present in the proper location to be inhibited by luminal flavonoids; and that the potent inhibitor, quercetin, was not itself transported by GLUT2. Our findings provide the essential foundation for a pilot clinical study to move forward, and this can now proceed.